Young women are in urgent need of female-controlled HIV prevention strategies, including antiretroviral (ARV) drugs for topical or oral pre-exposure prophylaxis (PrEP). Two daily oral ARV PrEP studies and one vaginal gel study demonstrated efficacy in preventing HIV-1 infection in women. However, two other studies of daily oral PrEP and one of daily tenofovir (TFV) gel were stopped early for futility. Prior studies suggest differences in ARV metabolism and cervical immune cell populations between otherwise similar U.S. and sub-Saharan African women, but specific differences in the genital mucosal environment that may impact ARV pharmacokinetics (PK), pharmacodynamics (PD) and efficacy are understudied. We hypothesize that young age, bacterial vaginosis (BV), semen exposure, and depot medroxyprogesterone acetate (DMPA) use are associated with increased activation of CD4-i- T cells, changes in soluble immunity, and loss of vaginal lactobacilli, which increase HIV acquisition risk and alter topical ARV PK/PD. In Project 3, mechanisms by which known HIV risks impact PK/PD will be investigated in four clinical studies: women will be studied at BV diagnosis and after successful treatment, in the setting of unprotected vaginal sexual intercourse, before and after initiation of DMPA, and by comparing sexually active adolescents to adult women. The ultimate goal is to utilize knowledge of the mechanisms underlying HIV acquisition risk to advance selection of effective, female-controlled PrEP strategies. We also propose a pre-Phase 1 tenofovir disoproxil fumarate (TDF) intravaginal ring (IVR) study in U.S. and African women at risk for HIV to assess the hypothesis that changes in mucosal immunity related to hormonal effects and/or changes in vaginal microbiota modulate PK/PD of TDF. Drug levels will be measured in plasma, genital tract secretions and epithelial tissue following 14 days of TDF ring use (Core B). Pyrosequencing and species-specific quantitative PCR assays will be performed to examine changes in vaginal microbiota that may modulate PK/PD of TDF. These studies will utilize innovative approaches to inform the development of topical ARV PrEP strategies and will investigate the key hypothesis that distinct populations of high risk women may require differential preventative strategies.